We hypothesize that exercise and caloric restriction inhibit carcinogenesis by common mechanisms and that crucial to both is the manner in which the body maintains energy homeostasis (energy balance) in the face of an imposed energy restricted state (ERS). We thus see convergence of these areas of investigation into one focus of research. In this grant application we propose to study the effects of ERS induced by exercise and caloric restriction to 1) investigate the origin(s) of the cancer inhibitory activity attributed to ERS, and 2) determine if ERS causes only a quantitative reduction in the occurrence of cancers or whether tumors with specific genetic alterations are selectively inhibited. This information will provide insights to the gene activities that are specifically being altered in ERS, ultimately shedding light on the mechanism of protection. Four specific aims are proposed. AIM 1. Does increased adrenal cortical activity play a specific role in the inhibition of mammary carcinogenesis by ERS? The hypothesis that elevated levels of cortical steroids play a causal role in the cancer inhibition due to energy restriction will be evaluated. Protein bound and free corticosteroids will be determined in plasma and urine by radioimmunoassay and specific metabolites detected by HPLC. AIM 2. Does ERS result in simply fewer carcinomas with the same pathogenetic markers or do the tumors that occur in energy restricted versus unrestricted rats represent pathogenetically distinct populations of tumors: Altered function of Ha-ras will be assessed using PCR-RFLP analyses and/or immunohistochemical techniques with quantification via videoimage analyses. Other pathogenetic markers will be investigated as their involvement in the tumor model is validated. AIM 3. Is the process of clonal expansion affected by ERS and if so, what mediates this effect: Expansion of cell populations bearing mutant Ha-ras alleles will be quantified via PCR. Alterations in clonal expansion will be further studied to determine if changes in cell proliferation or cell death are involved. AIM 4. Does ERS induce changes in the epithelial or mesenchymal component of the mammary gland that are characteristic of tissue remodeling and that may play a role in the observed inhibition of mammary carcinogenesis? ERS induces changes in the extracellular matrix of the mammary gland. Using Western and Northern blot techniques, the quantitative and qualitative nature of these changes will be identified and their relationship to inhibition of mammary carcinogenesis by ERS investigated. ERS is a profoundly effective, noninvasive, nonpharmacological approach to prevention of cancer in many organs, including the breast, yet the mechanism(s) by which it acts is unknown. Knowledge of its mechanism(s) may lead to novel, widely applicable and well tolerated approaches to cancer prevention and control for a major segment of the population.